For many years the medical profession has used clinical testing to determine total human serum calcium concentration in connection with diagnosis and treatment of various disease of the body including parathyroid disorders, tetany and bone structure diseases. Total calcium concentration of the serum includes both bound calcium and ionized or disassociated calcium. More recently, it has been recognized that the ionized calcium level is of significance in diagnosis and treatment of certain human malfunctions such as the diseases noted above. Knowing the total calcium concentration of a person, which can be measured by conventional means does not enable one to know the level of ionized calcium of that person. Thus, direct methods of measuring ionized calcium are desirable.
It has been difficult to clinically measure human serum ionized calcium concentrations of the body by conventional clinical laboratory procedures. However, one apparatus for directly determining ionized calcium concentration in protein-containing liquids such as blood serum has been developed and is the Orion Ionalyzer Serum Calcium Activity Flow-Through System such as model 99-20 (Orion Research, Inc., Cambridge, Mass. U.S.A.). This system, as is known, basically uses two or three aqueous standards of known ionized calcium concentration which are flowed in turn through a calcium electrode with the millivoltage produced being recorded. The millivoltage is plotted against the logarithm of the ionized calcium for each standard to produce a calibration graph. Blood serum sample millivoltage is then similarly measured and by means of the calibration graph its ionized calcium concentration extrapolated manually or automatically by an electrical calculator such as an Olivetti 101.
Thus, a method of directly measuring ionized calcium in a sample is available. However, certain problems arise in making useful and practical clinical determinations. When blood is withdrawn from its in vivo state in the body, its original pH in the body changes as time of exposure to air progresses. Thus, if ionized calcium is measured directly after withdrawal of the blood from a patient, reasonably accurate original or body serum ionized calcium concentration is obtained. However, such immediate measurement is often not practical because of distances between patients and the clinical laboratory as well as the volume of determinations to be made, shortages of personnel and the like.
The normal range of blood serum pH in humans is from about 7.27 to about 7.51 (at the 95% confidence limits) with a pH of 7.39 being considered an average, normal pH at normal body temperature of 37.degree.C.
It has now been found extremely important for certain clinical testing purposes to know the pH of blood serum, i.e., blood, when in the body and correct for this pH in a blood sample in order to determine the actual ionized calcium concentration of the blood serum, i.e., blood, when the sample was in the body. It has not been recognized that it is important to adjust for and make an ionized calcium determination on the sample by artificially varying the pH of the sample whereby exact original pH of the sample is accounted for in measurement if extremely accurate measurement is desired. Moreover, it has not heretofore been known that there is a straight line relationship between pH and ionized calcium concentration of serum when plotted on semi-log paper.